The emergence of bacterial strains resistant to most of the current antibacterial agents has a significant negative impact on public health. To combat bacterial resistance, new classes of antibacterial agents are needed that have expanded spectrum that includes the resistant strains. Particularly attractive are new classes of antibacterial drugs that inhibit novel bacterial targets. The potential benefits of these drugs are increased antibacterial potencies, expanded spectrum and significantly reduced frequencies for the emergence of new resistant strains. Rx3 Pharmaceuticals has developed a highly effective and experimentally facile method for MOA detection of antimicrobial hit compounds against the biowarfare pathogen B. anthracis. We have shown that we can apply this assay to hit compounds derived from high-throughput screens to determine their target specificity. Knowledge of the MOA of a hit compound enables SAR and SBDD efforts to improve potency and drug like properties. Quick identification of such lead-potential antimicrobial compounds within the MLSCN library will greatly accelerate the overall development of new antimicrobial drugs. Moreover, cross-correlation of these findings to information derived from other screens, such as mammalian cell toxicity, could greatly facilitate the prioritization of those compounds with the greatest potential to become the next clinically useful antibiotic against B. anthracis and other important bacterial pathogens. The goal of this work is identify novel leads for antibacterial drugs to address the rising threat of antibiotic- resistant infections and the potential threat of biowarfare bacterial pathogens. To accomplish this, we propose to screen the MLSCN compound collection for antibacterial compounds and then identify those compounds that act through a novel mechanism. [unreadable] [unreadable] [unreadable]